IDENTIFICATION AND CHARACTERIZATION OF THE CELL-SURFACE 70-KILODALTONSIALOGLYCOPROTEIN(S) AS A CANDIDATE RECEPTOR FOR ENCEPHALOMYOCARDITISVIRUS ON HUMAN NUCLEATED CELLS

The attachment of encephalomyocarditis (EMC) virus to human nucleated cells susceptible to virus infection was examined with HeLa and K562 c ell lines. Both cell types showed specific virus binding competitively blocked by unlabeled virions. The number of binding sites for EMC vir us on HeLa and K562 cells were approximately 1.6 x 10(5) and 3.5 x 10( 5) per cell, respectively, and dissociation binding constants were 1.1 and 2.7 nM, respectively. Treatment of cells with cycloheximide after pretreatment with trypsin eliminated EMC virus attachment, suggesting that the virus-binding moiety is proteinaceous in nature. Digestion o f cells, cell membranes, and sodium deoxycholate-solubilized cell memb ranes with proteases or neuraminidases or treatment of cells with lect ins demonstrated that the EMC virus-cell interaction is mediated by a sialoglycoprotein. Proteins with a molecular mass of 70 kDa were isola ted from detergent-solubilized cell membranes of both HeLa and K562 ce lls by EMC virus affinity chromatography. The purified proteins, as we ll as their 70-kDa-molecular-mass equivalents detected in intact surfa ce membranes of HeLa and K562 cells, specifically bound EMC virus in a virus overlay protein blot assay, whereas membranes from nonpermissiv e K562 D clone cells did not. Western immunoblot analysis with glycoph orin A-specific antibody confirmed that the identified 70-kDa binding site on K562 cells is not glycophorin A, which is the EMC virus recept or molecule on virus-nonpermissive human erythrocytes (HeLa cells do n ot express glycophorin A). These results indicate that EMC virus attac hment to permissive human cells is mediated by a cell surface sialogly coprotein(s) with a molecular mass of 70 kDa.